Pathogens of plants and animals produce effector proteins that are transferred into the cytoplasm of host cells to suppress host defenses. One type of plant pathogens, oomycetes, produces effector proteins with N-terminal RXLR and dEER motifs that enable entry into host cells. We show here that effectors of another pathogen type, fungi, contain functional variants of the RXLR motif, and that the oomycete and fungal RXLR motifs enable binding to the phospholipid, phosphatidylinositol-3-phosphate (PI3P). We find that PI3P is abundant on the outer surface of plant cell plasma membranes and, furthermore, on some animal cells. All effectors could also enter human cells, suggesting that PI3P-mediated effector entry may be very widespread in plant, animal and human pathogenesis. Entry into both plant and animal cells involves lipid raft-mediated endocytosis. Blocking PI3P binding inhibited effector entry, suggesting new therapeutic avenues.

In preparing the figures for this article for publication, we inadvertently assembled duplicated images of three panels in Figure 2 (Figure 2O, panel 2 was a duplicate of Figure 2N, panel 2; Figure 2R, panel 2 was a duplicate of Figure 2Q, panel 2; and Figure 2N, panel 3 was a duplicate of Figure 2K, panel 3). Also, in Figure S4, the leaf pictured in panel 8 of Figure S4A was mistakenly a picture of the same leaf as shown in panel 6. In the first two cases, the figures were correct in the original reviewed submission, and the errors occurred in revising the paper for resubmission. Corrected Figures 2 and S4 are shown below. The new figures do not alter any of the conclusions of the study. Finally, in the Experimental Procedures, under Lipid Filter-Binding Assays, the statement, ''Lipids were dissolved in DMSO then spotted .'' should read ''Lipids were dissolved in chloroform: methanol: water (65:30:8) then spotted .''The authors regret any confusion these errors may have caused. All errors have been corrected in the online version of the manuscript.

Gammaherpesviruses, including Epstein-Barr virus (EBV), Kaposi sarcoma-associated herpesvirus (KSHV, or HHV-8), and murine gammaherpesvirus 68 (MHV68, γHV68, or MuHV-4), are B cell-tropic pathogens that each encode at least 12 microRNAs (miRNAs). It is predicted that these regulatory RNAs facilitate infection by suppressing host target genes involved in a wide range of key cellular pathways. However, the precise contribution that gammaherpesvirus miRNAs make to viral life cycle and pathogenesis in vivo is unknown. MHV68 infection of mice provides a highly useful system to dissect the function of specific viral elements in the context of both asymptomatic infection and disease. Here, we report (i) analysis of in vitro and in vivo MHV68 miRNA expression, (ii) generation of an MHV68 miRNA mutant with reduced expression of all 14 pre-miRNA stem-loops, and (iii) comprehensive phenotypic characterization of the miRNA mutant virus in vivo. The profile of MHV68 miRNAs detected in infected cell lines varied with cell type and did not fully recapitulate the profile from cells latently infected in vivo. The miRNA mutant virus, MHV68.Zt6, underwent normal lytic replication in vitro and in vivo, demonstrating that the MHV68 miRNAs are dispensable for acute replication. During chronic infection, MHV68.Zt6 was attenuated for latency establishment, including a specific defect in memory B cells. Finally, MHV68.Zt6 displayed a striking attenuation in the development of lethal pneumonia in mice deficient in IFN-γ. These data indicate that the MHV68 miRNAs may facilitate virus-driven maturation of infected B cells and implicate the miRNAs as a critical determinant of gammaherpesvirus-associated disease.

Noncoding RNAs (ncRNAs) represent an intriguing and diverse class of molecules that are now recognized for their participation in a wide array of cellular processes. Viruses from multiple families have evolved to encode their own such regulatory RNAs; however, the specific in vivo functions of these ncRNAs are largely unknown. Epstein-Barr virus (EBV) and Kaposi’s sarcoma-associated herpesvirus (KSHV) are ubiquitous human pathogens that are associated with the development of numerous malignancies. Like EBV and KSHV, murine gammaherpesvirus 68 (MHV68) establishes lifelong latency in B cells and is associated with lymphomagenesis. The work described here reveals that the MHV68 ncRNA TMER4 acts at a critical bottleneck in local lymph nodes to facilitate hematogenous dissemination of the virus and establishment of latency at peripheral sites.

Serpins (serine protease inhibitors) are ubiquitous, complex, and highly active regulatory molecules that effectively control multiple coagulation, inflammatory, and neuroendocrine pathways (1-3). The amino acid sequence in the reactive center loop (RCL) 3 of serpins acts as bait for target serine proteases, initiating structural changes in the serpin-protease complex and culminating in suicide inhibition (1-3). This same RCL can insert into the neighboring ␤-sheet A in other serpins in serpinopathies, causing serpin aggregates induced by genetic mutations and causing disease as for anti-thrombin III (SERPIN C1, ATIII), ␣-1 antitrypsin (SERPIN A1, AAT), and neuroserpin (SERPIN I1, NSP). Whereas the amino acid residues in the RCL provide target P1-P1Ј sequences, referred to as a scissile bond, serpins also require the greater part of the protein structure to function with true serpin-protease inhibitory activity (4, 5). However, as for other proteins, peptides derived during protein metabolism may act to extend serpin activity beyond the initial suicide-inhibitory function, both increasing and decreasing responses (6 -9). In prior work, significant and prolonged antiinflammatory functions have been detected with myxomavirus-derived Serp-1 (10 -17) and mammalian serpin NSP purified protein injections in animal models of vascular disease (18,19). We have hypothesized that peptides produced by protease cleavage of the RCL sequence during natural proteolytic metabolism of Serp-1 or NSP may extend serpin activity, increasing anti-inflammatory activity after serpin-protease complex formation. Thus, these serpin RCL peptide metabolites have the theoretical potential to interfere with either protease activity by acting as a protease bait or inhibitors or to inhibit other serpins by inserting into the ␤-sheet.Many proteins have active metabolites providing additional and/or expanded functions. Peptides derived from calreticulin (20) and apolipoprotein E (Ep1B) (21) have proven anti-atherogenic activity, reducing inflammation and plaque growth in animal models. Serpins also have reported active terminal peptide metabolites. Among the serpins, angiotensinogen is a protein with serpin structure but lacking serpin inhibitory activity (6).